Project Summary Mitochondria are essential for energy production, but, if damaged, they become a major source of reactive oxygen species and proapoptotic factors. Increasing evidence suggests that mitochondrial dysfunction is a central event in the development of diabetic cardiomyopathy. However, the molecular mechanisms responsible for diabetes-caused mitochondrial dysfunction in cadiomyocytes remain poorly characterized. Our exciting preliminary data included in this application show that the levels of the FUN14 domain containing 1 (FUNDC1) protein, a mitochondrial outer-membrane protein, are highly increased in diabetic hearts resulting in mitochondrial dysfunction and diabetic cardiomyopathy. Mechanistically, aberrant FUNDC1 expression in diabetes increases endoplamsmic reticulum (ER)- mitochondria contact, which promotes Ca2+ transfer from the ER to the mitochondria and thus results in mitochondrial Ca2+ overload, cardiomyocyte death, and cardiac dysfunction. Cardiac-specific deletion of FUNDC1 reduces ER-mitochondria contacts, attenuating cardiac dysfunction in Type I and Type II diabetic mice. Thus, the central hypothesis of this proposal is that aberrant FUNDC1 expression in diabetes leads to cardiomyopathy by impairing mitochondrial function through enhancement of ER-mitochondria contacts. This hypothesis will be tested by using gain-/loss-of function and pharmacologic/genetic strategies in both animal models and cultured cardiomyocytes. Aim 1 is to establish the essential roles of increased FUNDC1 expression in the development of diabetic cardiomyopathy. In this Aim, we will test the hypotheis that enhanced FUNDC1 expression causes cardiac structural damage and dysfunction by compromising mitochondrial function in diabetes. Aim 2 is to elucidate the mechanism by which FUNDC1 upregulation in diabetes impairs mitochondrial function, leading to cardiomyopathy. In this aim, we will test the hypothesis that diabetes-enhanced FUNDC1 expression impairs mitochondrial function by promoting the ER- mitochondria contacts. We will determine if FUNDC1 mediates ER-mitochondria contacts in diabetic hearts, investigate whether increased FUNDC1 causes mitochondrial dysfunction and cardiomyocyte death by increasing Ca2+ transfer from ER to mitochondria, and examine whether diminishing ER- mitochondrial Ca2+ flux improves mitochondrial and cardiac function in diabetic hearts using IP3R2 cardiomyocyte-specific knockout mice. The completion of this highly innovative proposal will help develop a new paradigm for treating diabetic cardiomyopathy.